The present invention relates in general to the field of antimicrobial agents, and more particularly, to the characterization and isolation of agents that are responsible for antimicrobial activity of Aloe vera and its gel.
Without limiting the scope of the invention, its background is described in connection with the identification of novel anti-microbial agents isolated from Aloe vera, as an example.
Heretofore, in this field, organisms that cause infectious disease, namely, viruses, bacteria, fungi and multicellular parasites, humankind has sought to control their morbidity and mortality. With the isolation and characterization of powerful antibiotics, beginning over half a century ago, the balance of power between humans and microbes has been shifted toward humankind. For several decades after the introduction of penicillin in the 1940""s, for example, the conquest of infectious disease appeared imminent. The widespread use of antibiotics, added to the evolutionary flexibility of microbes, has made that victory less than certain.
An increasing number of bacteria, fungi and other microbes are developing resistance to antibiotics. A number of factors have contributed to the increase in microbes that are resistant to antimicrobial agents. The use of combinations of anti-microbial agents to treat nosocomial infections, particularly among patients whose immune systems are compromised by AIDS, chemotherapy, or immunosuppressive drugs, has led to a dramatic increase in multiple drug resistant (MDR) infections. Unfortunately, the future does not look bright in the war against infectious disease, as MDR strains of microbes continue to proceeding at an alarming rate. In fact, MDR strains are adapting faster than the introduction of new, more potent antibiotics.
Microbes have been developing strategies to cope with change for hundreds of millions of generations. In fact, some bacteria have generation cycles of 20 minutes, with each cycle providing the opportunity to evolve and adapt. Bacteria have adapted to an extraordinary range of conditions and developed defenses against all sorts of environmental threats, environmental and artificial. To a microbe the human body is just another environment to colonize. While antibiotics are just another toxic environmental agent against which the microbe must develop an escape strategy. For organisms with populations that have already adapted to such extreme environments as boiling underwater hot-springs, learning to cope and evade antibiotics was only a matter of time and evolution.
Overuse of antibiotics has contributed to the problem of MDR microbial strains. The indiscriminate use of antibiotics throughout the world contributes to the continued emergence of MDR strains of bacteria such as Pseudomonas, Streptococcus and Staphylococcus. MDR strains have evolved in large part because many patients fail to complete the required course of antibiotic treatment, allowing stronger members of the microbial pool to be selected for in the next round of treatment. Increases in ear and sinus infections in children have been caused by the use of antibiotics to treat viral infections, infections that are not susceptible to antibiotic treatment. The current trend in medicine is to prescribe second-line and even last-resort antibiotics in place of first-line antibiotics. Even when there is no reason to suspect resistance to first-line antibiotics, the drive toward using stronger, faster drugs is inevitable when faced with a sick patient. In the case of recurrent lung infections in cystic fibrosis patients, physicians have had no choice but to escalate to antibiotic treatment with second-line antibiotics, eventually causing the infecting bacteria to become resistant to all available antibiotics.
The emergence of MDR microbes has changed the balance between host and parasite, from a position in which the medical community seemed poised to achieve a conquest has lost ground in achieving a permanent conquest of microbial infection. But much has been learned in the process. Using a deeper understanding of microbes and their mechanisms of resistance, the biomedical community can continue to mount a broad array of defenses against them. The microbes growing resistance to traditional antibiotics has renewed the attention to medical basics, such as public health measures, that include a renewed effort to stem infectious diseases by increasing hygiene. For example, in HIV-infected populations, which have become breeding grounds for resistant microbes, renewed educational outreach efforts focus on the use of prophylactics.
Nosocomial infections present the greatest threat to immuno-compromised patients, because MDR microbes infect the most vulnerable patients. It is the increase in MDR of microbes, and in particular bacteria, that has led to a resurgence of interest in revitalizing and improving basic techniques (like hand-washing) for preventing the spread of infection. It has also increased the need for alternative, next-generation, anti-microbial agents. These antimicrobial agents, viz., anti-viral, anti-bacterial, anti-fungal and anti-parasitic, must also be safe for use in humans and other animals.
Antimicrobial-drug resistance is an increasingly important factor and poses a serious international challenge to public health in community and institutional settings. The list of resistant bacteria of major public health importance includes those causing tuberculosis, gonorrhea, pneumococcal infections, and hospital-acquired enterococcal and staphylococcal infections. Antimicrobial-drug resistance has resulted in prolonged and more serious illness, the use of more expensive and often more toxic drugs and drug combinations, and increased fatality rates.
While pharmaceutical and biotechnology companies are constantly developing novel products based on presently known antibiotics to overcome resistance, the next-generation of anti-microbial agents must break from the known approaches to isolate and characterize these activities. A better understanding of the microbiology and molecular genetics of microbial resistance is leading to the development of a new generation of anti-microbial agents that use an approach that is intended to attack standard mechanisms of action to kill bacteria or fungi. These so called new approaches to fighting microbial infections rely on variations of existing drugs having longer half-lives and more potent effects, but rely on the existing database of pharmaceutics to attempt to outpace the microbes ability to evolve.
Both competition for nutrients and bacteriocin production play a role in determining the establishment of microbial communities in nature. When analyzing symbiotic associations this may be further influenced by the presence of antimicrobial chemicals produced by the host. In the case of Aloe vera barbedensis, the plant has been shown for centuries to exert broad spectrum healing activities. The source of the antimicrobial agents isolated herein were determined, as were the distinct populations of bacteria, and their dynamics within the indigenous microflora of Aloe vera. Localization of specific microbial populations was assessed using both direct culture of dissected plant material and immunological detection within tissue sections. Relative size and population diversity were determined through direct culture. Immunological detection demonstrated discrete populations within specific plant structures.
Bacterial identification was accomplished using standard staining and biochemical analysis. To more completely identify the specific species of bacterium isolated and their role in the antibacterial activity isolated herein, the environmental specimens were further analyzed using restriction fragment length polymorphism (RFLP) analysis. RFLP analysis was used to differentiate the various species of Bacillus found within the plant as well as definitive identification of Aeromicrobioum species and Curtobacterium species.
The efficacy of aloe liquid (see e.g., Coats, Aloe Vera: The Inside Story) as an antimicrobial agent is shown herein to have a wide range of gram negative and gram positive bacteria. The antimicrobial agents of the present invention are shown herein to effectively kill, or greatly reduce or eliminate the growth rate of the following bacteria: Staphylococcus aureus, Streptococcus pneumonia, Streptococcus pyogenes, Pseudomonas aeruginosa, E. coli, Propionibacterium acne, Helicobacter pyloni, and Salmonella typhi. 
The anti-bacteriocidal activity demonstrated herein is not due to the preservatives used in the preparation of the clear gel or the isolation of the antibacterial components, as the antimicrobial agents isolated herein were isolatable from liquid collected directly from freshly cut whole leaves prior to used in the same killing assays.
In addition to the antimicrobial activities of its liquid, it also has been shown to be nontoxic even when taken internally. These properties combined provide the impetus for the use of the antimicrobial agents isolated herein when used alone or in combination. The present invention demonstrates the isolation and identification of new, non-toxic, FDA approved antimicrobial agents that have been identified and isolated from the Aloe plant. These agents are efficacious and nontoxic, with a broad spectrum of antimicrobial properties.
Generally, and in one form of the invention, a composition isolated from the gel liquid of Aloe vera including, at least one antimicrobial agent isolated from the clear gel isolated from the whole leaf of Aloe vera, wherein the antimicrobial agent is an agent produced by the Aloe vera or indigenous bacteria that colonize the Aloe vera, is disclosed.
Furthermore, a method of decreasing the growth of a broad spectrum of bacteria including the steps of, isolating at least one antibacterial agents from the clear gel of an aloe vera plant and directly contacting the bacteria with at least one antibacterial agent from aloe vera, is also disclosed.
The present invention is based on the recognition that aloe vera isolated have been used to treat, and increase the healing rate of, wounds and other infectious diseases. The present invention is also based on the recognition that antibacterial agents secreted by aloe vera and the bacteria that grow in the gel and rind of aloe vera and which exhibit a wide range antimicrobial activity when exposed directly to the target microbe can be isolated. The antibacterial agents isolated herein have molecular weights of about: 555,000; 470,000; 240,000; 160,000; 25,000 and 4,000 Daltons.
Also, the antibacterial agents isolated herein, are partially secreted by bacteria that grow in the gel and rind of Aloe vera. The secreted products of these species of bacteria exhibit a wide range antimicrobial activity when exposed directly to target microbes and each other and include: Aerobacterium, Bacillus, Curtobacterium, Arthrobacter, Sporosarcina, and Clavibacter.